## `summarise()` ungrouping output (override with `.groups` argument)
## [1] 4376
## `summarise()` ungrouping output (override with `.groups` argument)
## [1] 718
## `summarise()` ungrouping output (override with `.groups` argument)
## [1] 3856
## `summarise()` ungrouping output (override with `.groups` argument)
## [1] 584
## `summarise()` regrouping output by 'sample', 'age', 'gender', 'collection_date', 'week', 'year', 'lineage', 'HOCI', 'HCW', 'protocol', 'mapped_reads', 'ect', 'rct' (override with `.groups` argument)

Main Figures

Figure 1

Sub-genomic RNA Expression is Increased in B.1.1.7 infections

## `summarise()` ungrouping output (override with `.groups` argument)
## # A tibble: 5 x 10
##   orf   .y.   group1 group2    n1    n2 statistic        p    p.adj p.adj.signif
##   <fct> <chr> <chr>  <chr>  <int> <int>     <dbl>    <dbl>    <dbl> <chr>       
## 1 S     total B.1.1… B.1.1…   764   729   125745  3.84e-75 1.92e-74 ****        
## 2 E     total B.1.1… B.1.1…   764   729   217058  1.63e-13 4.89e-13 ****        
## 3 M     total B.1.1… B.1.1…   764   729   255940. 7.00e- 3 7.00e- 3 **          
## 4 N     total B.1.1… B.1.1…   764   729   127596  2.25e-73 9.00e-73 ****        
## 5 ORF6  total B.1.1… B.1.1…   764   729   244833  5.34e- 5 1.07e- 4 ****
## # A tibble: 5 x 8
##   .y.   group1  group2  effsize orf      n1    n2 magnitude
## * <chr> <chr>   <chr>     <dbl> <fct> <int> <int> <ord>    
## 1 total B.1.177 B.1.1.7  0.475  S       764   729 moderate 
## 2 total B.1.177 B.1.1.7  0.191  E       764   729 small    
## 3 total B.1.177 B.1.1.7  0.0700 M       764   729 small    
## 4 total B.1.177 B.1.1.7  0.469  N       764   729 moderate 
## 5 total B.1.177 B.1.1.7  0.105  ORF6    764   729 small
## # A tibble: 5 x 10
##   orf   .y.   group1 group2    n1    n2 statistic        p    p.adj p.adj.signif
##   <fct> <chr> <chr>  <chr>  <int> <int>     <dbl>    <dbl>    <dbl> <chr>       
## 1 S     total B.1.1… B.1.1…   179   150     5534. 4.18e-20 1.67e-19 ****        
## 2 E     total B.1.1… B.1.1…   179   150    11342. 1.50e- 2 1.60e- 2 *           
## 3 M     total B.1.1… B.1.1…   179   150    15702. 8.00e- 3 1.60e- 2 **          
## 4 N     total B.1.1… B.1.1…   179   150     4352  4.66e-26 2.33e-25 ****        
## 5 ORF6  total B.1.1… B.1.1…   179   150    10121  1.21e- 4 3.63e- 4 ***
## # A tibble: 5 x 10
##   orf   .y.   group1 group2    n1    n2 statistic        p    p.adj p.adj.signif
##   <fct> <chr> <chr>  <chr>  <int> <int>     <dbl>    <dbl>    <dbl> <chr>       
## 1 S     sgRN… B.1.1… B.1.1…   204   138     6900. 1.26e-15 6.30e-15 ****        
## 2 E     sgRN… B.1.1… B.1.1…   204   138    12444. 6.90e- 2 1.38e- 1 ns          
## 3 M     sgRN… B.1.1… B.1.1…   204   138    11952. 1.80e- 2 5.40e- 2 *           
## 4 N     sgRN… B.1.1… B.1.1…   204   138     7498. 2.25e-13 9.00e-13 ****        
## 5 ORF6  sgRN… B.1.1… B.1.1…   204   138    13168. 3.11e- 1 3.11e- 1 ns
## `geom_smooth()` using formula 'y ~ x'
## `geom_smooth()` using formula 'y ~ x'

Figure 2

Relationship between day of symptom onset at sampling and sub-genomic RNA levels in B.1.1.7 and B.1.177 infections

## `summarise()` regrouping output by 'sample', 'lineage', 'Day_of_symptoms' (override with `.groups` argument)
## `summarise()` ungrouping output (override with `.groups` argument)
## 
## Call:
## lm(formula = sgRPTg_HQ + sgRPTg_LQ ~ lineage + Day_of_symptoms, 
##     data = for_model_s)
## 
## Residuals:
##    Min     1Q Median     3Q    Max 
## -363.7  -98.9  -51.6    4.8 6370.6 
## 
## Coefficients:
##                 Estimate Std. Error t value Pr(>|t|)    
## (Intercept)       67.795     32.832   2.065   0.0392 *  
## lineageB.1.1      -5.401     38.010  -0.142   0.8870    
## lineageB.1.1.7   295.879     37.450   7.901 7.64e-15 ***
## lineageB.1.177    50.925     36.341   1.401   0.1614    
## Day_of_symptoms    6.480      3.582   1.809   0.0707 .  
## ---
## Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1
## 
## Residual standard error: 344.6 on 951 degrees of freedom
## Multiple R-squared:  0.1146, Adjusted R-squared:  0.1109 
## F-statistic: 30.79 on 4 and 951 DF,  p-value: < 2.2e-16
## 
## Call:
## lm(formula = sgRPTg_HQ + sgRPTg_LQ ~ lineage + Day_of_symptoms, 
##     data = for_model_s)
## 
## Residuals:
##    Min     1Q Median     3Q    Max 
## -363.7  -98.9  -51.6    4.8 6370.6 
## 
## Coefficients:
##                 Estimate Std. Error t value Pr(>|t|)    
## (Intercept)       67.795     32.832   2.065   0.0392 *  
## lineageB.1.1      -5.401     38.010  -0.142   0.8870    
## lineageB.1.1.7   295.879     37.450   7.901 7.64e-15 ***
## lineageB.1.177    50.925     36.341   1.401   0.1614    
## Day_of_symptoms    6.480      3.582   1.809   0.0707 .  
## ---
## Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1
## 
## Residual standard error: 344.6 on 951 degrees of freedom
## Multiple R-squared:  0.1146, Adjusted R-squared:  0.1109 
## F-statistic: 30.79 on 4 and 951 DF,  p-value: < 2.2e-16
## 
## Call:
## lm(formula = sgRPTg_HQ + sgRPTg_LQ ~ lineage + Day_of_symptoms, 
##     data = for_model_m)
## 
## Residuals:
##    Min     1Q Median     3Q    Max 
## -734.0 -269.0  -62.2   61.2 3270.4 
## 
## Coefficients:
##                 Estimate Std. Error t value Pr(>|t|)    
## (Intercept)       78.083     46.293   1.687    0.092 .  
## lineageB.1.1      35.578     53.593   0.664    0.507    
## lineageB.1.1.7   605.706     52.803  11.471   <2e-16 ***
## lineageB.1.177   582.735     51.239  11.373   <2e-16 ***
## Day_of_symptoms    7.961      5.050   1.576    0.115    
## ---
## Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1
## 
## Residual standard error: 485.9 on 951 degrees of freedom
## Multiple R-squared:  0.2402, Adjusted R-squared:  0.237 
## F-statistic: 75.16 on 4 and 951 DF,  p-value: < 2.2e-16
## 
## Call:
## lm(formula = sgRPTg_HQ + sgRPTg_LQ ~ lineage + Day_of_symptoms, 
##     data = for_model_n)
## 
## Residuals:
##     Min      1Q  Median      3Q     Max 
##  -473.9   -98.1   -37.5    44.1 12263.4 
## 
## Coefficients:
##                 Estimate Std. Error t value Pr(>|t|)    
## (Intercept)       77.317     42.032   1.839  0.06616 .  
## lineageB.1.1      40.524     48.661   0.833  0.40517    
## lineageB.1.1.7   420.032     47.944   8.761  < 2e-16 ***
## lineageB.1.177   149.509     46.523   3.214  0.00135 ** 
## Day_of_symptoms    2.427      4.586   0.529  0.59680    
## ---
## Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1
## 
## Residual standard error: 441.1 on 951 degrees of freedom
## Multiple R-squared:  0.1173, Adjusted R-squared:  0.1136 
## F-statistic:  31.6 on 4 and 951 DF,  p-value: < 2.2e-16
## 
## Call:
## lm(formula = sgRPTg_HQ + sgRPTg_LQ ~ lineage + Day_of_symptoms, 
##     data = for_model_orf6)
## 
## Residuals:
##    Min     1Q Median     3Q    Max 
## -276.4  -89.1  -54.0   -7.5 5705.6 
## 
## Coefficients:
##                 Estimate Std. Error t value Pr(>|t|)   
## (Intercept)       77.566     29.817   2.601  0.00943 **
## lineageB.1.1      36.815     34.519   1.067  0.28646   
## lineageB.1.1.7    -9.326     34.010  -0.274  0.78398   
## lineageB.1.177   -10.737     33.003  -0.325  0.74501   
## Day_of_symptoms    9.720      3.253   2.988  0.00288 **
## ---
## Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1
## 
## Residual standard error: 312.9 on 951 degrees of freedom
## Multiple R-squared:  0.01596,    Adjusted R-squared:  0.01182 
## F-statistic: 3.856 on 4 and 951 DF,  p-value: 0.004092
SEMNORF6
(Intercept)67.79 *  67.79 *  78.08    77.32    77.57 **
(32.83)   (32.83)   (46.29)   (42.03)   (29.82)  
lineageB.1.1-5.40    -5.40    35.58    40.52    36.81   
(38.01)   (38.01)   (53.59)   (48.66)   (34.52)  
lineageB.1.1.7295.88 ***295.88 ***605.71 ***420.03 ***-9.33   
(37.45)   (37.45)   (52.80)   (47.94)   (34.01)  
lineageB.1.17750.93    50.93    582.73 ***149.51 ** -10.74   
(36.34)   (36.34)   (51.24)   (46.52)   (33.00)  
Day_of_symptoms6.48    6.48    7.96    2.43    9.72 **
(3.58)   (3.58)   (5.05)   (4.59)   (3.25)  
N956       956       956       956       956      
R20.11    0.11    0.24    0.12    0.02   
*** p < 0.001; ** p < 0.01; * p < 0.05.
## Loading required namespace: broom.mixed
## Loading required namespace: broom.mixed
## Loading required namespace: broom.mixed
## Loading required namespace: broom.mixed
## Loading required namespace: broom.mixed
## Picking joint bandwidth of 0.895
## `geom_smooth()` using method = 'loess' and formula 'y ~ x'

Figure 3

A Non-canonical sgRNA representing ORF9b is highly expressed in B.1.1.7 due to a triple nucleotide mutation in nucleocapsid leading to the D3L substitution.

## `summarise()` regrouping output by 'start', 'y', 'lineage' (override with `.groups` argument)
## # A tibble: 1 x 7
##   .y.   group1 group2  effsize    n1    n2 magnitude
## * <chr> <chr>  <chr>     <dbl> <int> <int> <ord>    
## 1 total Other  B.1.1.7   0.761   288   535 large

Supplementary Figures

Figure S2

Normalized sgRNA for ORF3a, ORF7a, ORF8 and ORF10

## # A tibble: 5 x 9
##   orf   .y.   group1  group2     n1    n2 statistic         p     p.adj
##   <fct> <chr> <chr>   <chr>   <int> <int>     <dbl>     <dbl>     <dbl>
## 1 N*    total B.1.177 B.1.1.7   764   729    46262. 1.15e-212 5.75e-212
## 2 ORF10 total B.1.177 B.1.1.7   764   729   275128  1.30e-  1 1.30e-  1
## 3 ORF3a total B.1.177 B.1.1.7   764   729   298928  1.08e-  2 2.16e-  2
## 4 ORF7a total B.1.177 B.1.1.7   764   729   313123  6.51e-  6 2.60e-  5
## 5 ORF8  total B.1.177 B.1.1.7   764   729   294984  1.51e-  3 4.53e-  3
## # A tibble: 5 x 9
##   orf   .y.   group1  group2     n1    n2 statistic        p    p.adj
##   <fct> <chr> <chr>   <chr>   <int> <int>     <dbl>    <dbl>    <dbl>
## 1 N*    total B.1.177 B.1.1.7   179   150     1443  3.98e-55 1.99e-54
## 2 ORF10 total B.1.177 B.1.1.7   179   150    13350  7.71e- 1 1.00e+ 0
## 3 ORF3a total B.1.177 B.1.1.7   179   150    11236  6.58e- 3 2.63e- 2
## 4 ORF7a total B.1.177 B.1.1.7   179   150    13374. 9.49e- 1 1.00e+ 0
## 5 ORF8  total B.1.177 B.1.1.7   179   150    12865  2.80e- 1 8.40e- 1

Figure S3

Comparison of Mapped Reads and Genomic RNA (i.e. non-sgRNA) reads between B.1.177 and B.1.1.7 infections.

## # A tibble: 1 x 7
##   .y.          group1  group2  effsize    n1    n2 magnitude
## * <chr>        <chr>   <chr>     <dbl> <int> <int> <ord>    
## 1 mapped_reads B.1.177 B.1.1.7  0.0810   943   879 small
## # A tibble: 1 x 7
##   .y.     group1  group2  effsize    n1    n2 magnitude
## * <chr>   <chr>   <chr>     <dbl> <int> <int> <ord>    
## 1 total_g B.1.177 B.1.1.7   0.154   943   879 small
## # A tibble: 1 x 7
##   .y.   group1  group2  effsize    n1    n2 magnitude
## * <chr> <chr>   <chr>     <dbl> <int> <int> <ord>    
## 1 total B.1.177 B.1.1.7   0.285   943   879 small

Figure S4

Linear regression model output evaluating impact of lineage and Day of symptoms at sampling on ORF3a, ORF7a, ORF8 and ORF10.

N*ORF10ORF3aORF7aORF8
(Intercept)-0.10    0.01 1.92 3.34 0.85 *
(0.56)   (0.02)(3.40)(14.16)(0.43) 
lineageB.1.12.24 ***0.00 6.81 1.64 -0.79  
(0.64)   (0.02)(3.94)(16.42)(0.50) 
lineageB.1.1.73.95 ***0.03 -0.11 16.55 -0.35  
(0.63)   (0.02)(3.88)(16.17)(0.49) 
lineageB.1.1770.05    0.01 0.56 -0.18 -0.43  
(0.62)   (0.02)(3.76)(15.69)(0.48) 
Day_of_symptoms0.03    -0.00 -0.28 0.34 0.03  
(0.06)   (0.00)(0.37)(1.55)(0.05) 
N957       957    957    957    957     
R20.08    0.00 0.01 0.00 0.00  
*** p < 0.001; ** p < 0.01; * p < 0.05.
## Loading required namespace: broom.mixed
## Loading required namespace: broom.mixed
## Loading required namespace: broom.mixed
## Loading required namespace: broom.mixed
## Loading required namespace: broom.mixed

Figure S5

Normalized sgRNA Expression in Healthcare Workers infected with B.1.177 and B.1.1.7 Lineages.

Figure S6

Normalized sgRNA Expression in B.1.1.7 Stratified by Sex

Figure S7

Normalized sgRNA Expression in B.1.1.7 Stratified by Age

Figure S8

Normalized sgRNA expression stratified by days since symptom onset for ORF3a, ORF7a, ORF8 and ORF10.